HRI depletion cooperates with pharmacologic inducers to elevate fetal hemoglobin and reduce sickle cell formation
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ABSTRACT:
PROVIDER: PRJNA631789 | ENA |
REPOSITORIES: ENA
Dataset's files
| Action | DRS | |||
|---|---|---|---|---|
| SRR11773356_1.fastq.gz | Fastqsanger.gz | |||
| SRR11773356_2.fastq.gz | Fastqsanger.gz | |||
| SRR11773357_1.fastq.gz | Fastqsanger.gz | |||
| SRR11773357_2.fastq.gz | Fastqsanger.gz | |||
| SRR11773358_1.fastq.gz | Fastqsanger.gz |
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Project description:Increasing fetal hemoglobin (HbF) provides clinical benefit in patients with sickle cell disease (SCD). We recently identified heme-regulated inhibitor (HRI, EIF2AK1) as a novel HbF regulator. Since HRI is an erythroid-specific protein kinase it presents a potential target for pharmacologic intervention. We find that maximal HbF induction requires >80-85% HRI depletion. As it remains unclear whether this degree of HRI inhibition can be achieved pharmacologically, we explored whether HRI knockdown can be combined with pharmacologic HbF inducers to achieve greater HbF production and minimize potential adverse effects associated with treatments. Strongly cooperative HbF induction was observed when HRI depletion was combined with exposure to pomalidomide or the EHMT1/2 inhibitor UNC0638, but not hydroxyurea. Mechanistically, reduction in the levels of the HbF repressor BCL11A reflected the cooperativity of HRI loss and pomalidomide treatment, whereas UNC0638 did not modulate BCL11A levels. In conjunction with HRI loss, pomalidomide maintained its HbF inducing activity at 10-fold lower concentrations, at which condition there were minimal observed detrimental effects on erythroid cell maturation and viability as well as fewer alterations in the erythroid transcriptome. In sum, this study provides a foundation for the exploration of combining future small molecule HRI inhibitors with additional pharmacologic HbF inducers to maximize HbF production and preserve erythroid cell functionality for the treatment of SCD and other hemoglobinopathies.
2020-09-21 | GSE150341 | GEO
Project description:Increasing fetal hemoglobin (HbF) provides clinical benefit in patients with sickle cell disease (SCD). We recently identified heme-regulated inhibitor (HRI, EIF2AK1), as a novel HbF regulator. Because HRI is an erythroid-specific protein kinase, it presents a potential target for pharmacologic intervention. We found that maximal HbF induction required >80% to 85% HRI depletion. Because it remains unclear whether this degree of HRI inhibition can be achieved pharmacologically, we explored whether HRI knockdown can be combined with pharmacologic HbF inducers to achieve greater HbF production and minimize potential adverse effects associated with treatments. Strongly cooperative HbF induction was observed when HRI depletion was combined with exposure to pomalidomide or the EHMT1/2 inhibitor UNC0638, but not to hydroxyurea. Mechanistically, reduction in the levels of the HbF repressor BCL11A reflected the cooperativity of HRI loss and pomalidomide treatment, whereas UNC0638 did not modulate BCL11A levels. In conjunction with HRI loss, pomalidomide maintained its HbF-inducing activity at 10-fold lower concentrations, in which condition there were minimal observed detrimental effects on erythroid cell maturation and viability, as well as fewer alterations in the erythroid transcriptome. When tested in cells from patients with SCD, combining HRI depletion with pomalidomide or UNC0638 achieved up to 50% to 60% HbF and 45% to 50% HbF, respectively, as measured by high-performance liquid chromatography, and markedly counteracted cell sickling. In summary, this study provides a foundation for the exploration of combining future small-molecule HRI inhibitors with additional pharmacologic HbF inducers to maximize HbF production and preserve erythroid cell functionality for the treatment of SCD and other hemoglobinopathies.
| S-EPMC7509868 | biostudies-literature
Project description:Serratia sp. HRI Genome sequencing and assembly
| PRJNA580358 | ENA
Project description:Iron and heme play central roles in red blood cell production. However, the mechanisms by which iron and heme levels coordinate erythropoiesis remain incompletely understood. HRI is a heme-regulated kinase that controls translation by phosphorylating eIF2a. Here, we investigate the global impact of iron, heme and HRI on protein translation in vivo in murine primary erythroblasts using ribosome profiling. By defining the underlying changes in translation during iron and HRI deficiencies, we validate known regulators of this process, including Atf4, and identify novel pathways such as co-regulation of ribosomal protein mRNA translation. Surprisingly, we found that heme and HRI pathways, but not iron-regulated pathways, mediate the major protein translational and transcriptional responses to iron deficiency in erythroblasts in vivo and thereby identify previously unappreciated regulators of erythropoiesis. Our genome-wide study uncovers the major impact of the HRI-mediated integrated stress response for the adaptation to iron deficiency anemia.
2018-09-03 | GSE119365 | GEO
Project description:Increasing fetal hemoglobin (HbF) levels in adult red blood cells provides clinical benefit to patients with sickle cell disease and some forms of beta-thalassemia. To identify potentially druggable HbF regulators in adult human erythroid cells, we employed a protein kinase-domain focused CRISPR/Cas9-based genetic screen with a newly optimized sgRNA scaffold. The screen uncovered the heme-regulated inhibitor HRI (also known as EIF2AK1), an erythroid-specific kinase that controls protein translation, as an HbF repressor. HRI depletion markedly increased HbF production in a specific manner and reduced sickling in cultured erythroid cells. Diminished expression of the HbF repressor BCL11A accounted in large part for the effects of HRI depletion. Taken together, these results suggest HRI as a potential therapeutic target for hemoglobinopathies.
2018-07-20 | GSE115687 | GEO
Project description:To study the mitochondrial stress response HRI-eIF2a-ATF4 signaling pathway in mitochondrial cardiomyopathy , we generated Ptpmt1 cardiomyocytes-specific Knockout mcie, Eif2a mutant mcie, Gcn2 Knockout mcie, and Hri Knockout mcie
2022-04-21 | GSE201042 | GEO
Project description:Overcoming the silencing of the fetal γ-globin gene has been a long-standing goal in the treatment of sickle cell disease (SCD). The major transcriptional enhancer of the β-globin locus, called the locus control region (LCR), dynamically interacts with the developmental stage-appropriate β-type globin genes via chromatin looping, a process requiring the protein Ldb1. In adult erythroid cells, the LCR can be redirected from the adult β- to the fetal γ-globin promoter by tethering Ldb1 to the human γ-globin promoter with custom-designed zinc finger (ZF) proteins (ZF-Ldb1), leading to reactivation of γ-globin gene expression. To compare this approach to pharmacologic reactivation of fetal hemoglobin (HbF), hematopoietic cells from patients with SCD were treated with a lentivirus expressing the ZF-Ldb1 or with chemical HbF inducers. The HbF increase in cells treated with ZF-Ldb1 was more than double that observed with decitabine and pomalidomide; butyrate had an intermediate effect whereas tranylcypromine and hydroxyurea showed relatively low HbF reactivation. ZF-Ldb1 showed comparatively little toxicity, and reduced sickle hemoglobin (HbS) synthesis as well as sickling of SCD erythroid cells under hypoxic conditions. The efficacy and low cytotoxicity of lentiviral-mediated ZF-Ldb1 gene transfer compared with the drug regimens support its therapeutic potential for the treatment of SCD.
| S-EPMC5000846 | biostudies-other
Project description:Through a CRISPR-Cas9 guided loss-of-function screen in human erythroid cells we identified transcription factor ATF4, a known HRI-regulated protein, as a novel γ-globin repressor. ATF4 binds to a BCL11A enhancer to augment promoter contacts, stimulates BCL11A transcription to repress γ-globin expression. Notably, mice deficient for HRI displayed normal Bcl11a levels, suggesting species selective regulation that we explain here by demonstrating that the analogous ATF4 motif at the murine Bcl11a enhancer is largely dispensable. This illustrates potential limits of commonly used murine models of globin gene regulation.
2020-04-17 | GSE143961 | GEO
Project description:Through a CRISPR-Cas9 guided loss-of-function screen in human erythroid cells we identified transcription factor ATF4, a known HRI-regulated protein, as a novel γ-globin repressor. ATF4 binds to a BCL11A enhancer to augment promoter contacts, stimulates BCL11A transcription to repress γ-globin expression. Notably, mice deficient for HRI displayed normal Bcl11a levels, suggesting species selective regulation that we explain here by demonstrating that the analogous ATF4 motif at the murine Bcl11a enhancer is largely dispensable. This illustrates potential limits of commonly used murine models of globin gene regulation.
2020-04-17 | GSE143962 | GEO
Project description:The major heat shock protein Hsp70 has been shown to form a complex with a scaffold protein Bag3, linking it to multiple signaling pathways. Via these interactions, the Hsp70-Bag3 module functions as a proteotoxicity sensor that controls cell signaling. Here, as a tool to identify signaling pathways regulated by this complex, we utilized JG-98, an allosteric inhibitor of Hsp70 that blocks its interaction with Bag3. Gene expression profiling followed by the pathway analysis indicated that a set of signaling pathways including the unfolded protein response (UPR) was activated by JG-98. Surprisingly, only the translation initiation factor eIF2a-associated branch of the UPR was activated under these conditions, while other UPR branches mediating induction of ER chaperones were not induced, suggesting that the response was not related to ER proteotoxicity and thus to ER-associated kinase PERK1. Indeed, induction of the UPR genes under these conditions was dependent on activation of a distinct cytoplasmic eIF2a kinase, HRI. We demonstrated that the Hsp70-Bag3 complex directly interacted with HRI and regulated phosphorylation of eIF2a upon induction of cytoplasmic proteotoxicity. Therefore, we uncovered a novel signaling response, which regulates cell death upon the buildup of abnormal protein species in cytoplasm via an Hsp70-Bag3-HRI-eIF2a axis.
2021-04-14 | GSE171440 | GEO